Gene transfer for the treatment of cardiovascular diseases is bedeviled by inability to obtain, safely and easily, sufficient cardiac transgene expression. Current methods of gene transfer for heart disease include intramuscular injection into heart muscle or intracoronary delivery, approaches that are cumbersome to apply. Consequently, we have considered the usefulness of a vector encoding a paracrine-type transgene. In this approach, the transgene acts as a hormone, having cardiac effects after being released to the circulation from a distant site. This approach would circumvent the problem of attaining high yield cardiac gene transfer and enable patients to be treated by a systemic injection during an office visit. Furthermore, the approach proposed would eliminate the need for intravenous delivery of therapeutic peptides and thereby circumvent repeated and prolonged hospital stays, high morbidity, and enormous economic costs. The most suited vector to achieve these goals is the adeno-associated virus type 8 (AAV8), which provides long term and extensive expression after intravenous delivery in rodents, pigs, and primates. Urocortin-2, a recently discovered corticotrophin releasing factor family vasoactive peptide, acts via corticotropin-releasing factor type 2 receptors, which are robustly expressed in the heart and vasculature. Studies in animals and patients with congestive heart failure have shown favorable hemodynamic effects of urocortin-2 peptide infusions, including increased contractile function independent of loading, indicating direct cardiac effects. Urocortin-2 is an ideal selection as a therapeutic transgene in the proposed studies. We have established that intravenous delivery of AAV8 using the chicken -actin promoter provides sustained high serum levels of UCn2 and increases function of the normal and failing mouse heart. To develop and refine such an approach we propose to determine, in sequential studies in mice and pigs: a) the optimal system to provide regulated transgene expression to enable fine-tuning of plasma transgene levels, and allow turning expression off and on as needed; and b) the safety and efficacy of urocortin-2 gene transfer using this optimal, paracrine-based approach in a mouse model of CHF. Subsequently, in normal pigs we will determine: a) the minimally effective vector dose required to increase serum urocortin-2; b) biodistribution of the vector and transgene; and c) toxicity. HYPOTHESIS: Intravenous injection of an AAV8 vector with regulated expression of urocortin-2 will, through paracrine-mediated actions, have favorable effects on the failing heart. Aim 1. To determine the optimal regulated expression system to achieve long term and dose-responsive urocortin-2 expression in plasma, with minimal immune response and toxicity Aim 2. To test the optimal AAV8 vector providing regulated expression of urocortin-2 (identified in Aim 1) and determine in mice with and without heart failure: a) plasma levels over 12 months; b) efficacy for increasing function of the failing heart and reducing mortality; c) mechanisms for beneficial effects; and d) biodistribution and toxicity of the vector and transgene Aim 3. To test the optimal AAV vector encoding urocortin-2 (confirmed in Aim 2) in normal pigs to determine: a) plasma levels of urocortin-2; b) biodistribution of the vector and transgene; c) toxicity These mechanistic and proof-of-concept studies are designed to be sufficient in scope to lay the groundwork for future studies to be conducted in a CHF model in pigs and subsequently file an IND to initiate a clinical trial.